Upregulation of lipocalin-2 (LCN2) in osteoarthritic cartilage is not necessary for cartilage destruction in mice

Choi, W.S.,Chun, J.S. Published for the Society by Baillère Tinda 2017 OSTEOARTHRITIS AND CARTILAGE Vol.25 No.3

Objective: Lipocalin-2 (LCN2) is a recently characterized adipokine that is upregulated in chondrocytes treated with pro-inflammatory mediators and in the synovial fluid of osteoarthritis (OA) patients. Here, we explored the in vivo functions of LCN2 in OA cartilage destruction in mice. Methods: The expression levels of LCN2 were determined at the mRNA and protein levels in primary cultured mouse chondrocytes and in human and mouse OA cartilage. Experimental OA was induced in wild-type (WT) or Lcn2-knockout (KO) mice by destabilization of the medial meniscus (DMM) or intra-articular (IA) injection of adenoviruses expressing hypoxia-inducible factor (HIF)-2α (Ad-Epas1), ZIP8 (Ad-Zip8), or LCN2 (Ad-Lcn2). The effect of LCN2 overexpression on the cartilage of WT mice was examined by IA injection of Ad-Lcn2. Results: LCN2 mRNA levels in chondrocytes were markedly increased by the pro-inflammatory cytokines, interleukin (IL)-1β and tumor necrosis factor-α (TNF-α), and by previously identified catabolic regulators of OA, such as HIF-2α and components of the zinc-ZIP8-MTF1 axis. LCN2 protein levels were also markedly increased in human OA cartilage and cartilage from various experimental mouse models of OA. However, overexpression of LCN2 in chondrocytes did not modulate the expression of cartilage matrix molecules or matrix-degrading enzymes. Furthermore, LCN2 overexpression in mouse cartilage via IA injection of Ad-Lcn2 did not cause OA pathogenesis, and Lcn2 KO mice showed no alteration in DMM-induced OA cartilage destruction. Conclusions: Our observations collectively suggest that upregulation of LCN2 in OA cartilage is not sufficient or necessary for OA cartilage destruction in mice.

Regulation of senescence associated signaling mechanisms in chondrocytes for cartilage tissue regeneration

Ashraf, S.,Cha, B.H.,Kim, J.S.,Ahn, J.,Han, I.,Park, H.,Lee, S.H. Published for the Society by Baillère Tinda 2016 OSTEOARTHRITIS AND CARTILAGE Vol.24 No.2

<P>Adult articular chondrocytes undergo slow senescence and dedifferentiation during in vitro expansion, restricting successful cartilage regeneration. A complete understanding of the molecular signaling pathways involved in the senescence and dedifferentiation of chondrocytes is essential in order to better characterize chondrocytes for cartilage tissue engineering applications. During expansion, cell fate is determined by the change in expression of various genes in response to aspects of the microenvironment, including oxidative stress, mechanical stress, and unsuitable culture conditions. Rapid senescence or dedifferentiation not only results in the loss of the chondrocytic phenotype but also enhances production of inflammatory mediators and matrix-degrading enzymes. This review focuses on the two groups of genes that play direct and indirect roles in the induction of senescence and dedifferentiation. Numerous degenerative signaling pathways associated with these genes have been reported. Upregulation of the genes interleukin 1 beta (IL-1 beta), p53, p16, p21, and p38 mitogen-activated protein kinase (MAPK) is responsible for the direct induction of senescence, whereas downregulation of the genes transforming growth factor-beta (TGF-beta), bone morphogenetic protein-2 (BMP-2), SRY (sex determining region Y)-box 9 (SOX9), and insulin-like growth factor-1 (IGF-1), indirectly induces senescence. In senescent and dedifferentiated chondrocytes, it was found that TGF-beta, BMP-2, SOX9, and IGF-1 are downregulated, while the levels of IL-1 beta, p53, p16, p21, and p38 MAPK are upregulated followed by inhibition of the normal molecular functioning of the chondrocytes. This review helps to elucidate the underlying mechanism in degenerative cartilage disease, which may help to improve cartilage tissue regeneration techniques. (C) 2015 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.</P>

Reciprocal regulation by hypoxia-inducible factor-2α and the NAMPT-NAD⁺-SIRT axis in articular chondrocytes is involved in osteoarthritis

Oh, H.,Kwak, J.S.,Yang, S.,Gong, M.K.,Kim, J.H.,Rhee, J.,Kim, S.K.,Kim, H.E.,Ryu, J.H.,Chun, J.S. Published for the Society by Baillère Tinda 2015 Osteoarthritis and cartilage Vol.23 No.12

Objective: Hypoxia-inducible factor-2α (HIF-2α) transcriptionally upregulates Nampt in articular chondrocytes. NAMPT, which exhibits nicotinamide phosphoribosyltransferase activity, in turn causes osteoarthritis (OA) in mice by stimulating the expression of matrix-degrading enzymes. Here, we sought to elucidate whether HIF-2α activates the NAMPT-NAD<SUP>+</SUP>-SIRT axis in chondrocytes and thereby contributes to the pathogenesis of OA. Methods: Assays of NAD levels, SIRT activity, reporter gene activity, mRNA, and protein levels were conducted in primary cultured mouse articular chondrocytes. Experimental OA in mice was induced by intra-articular (IA) injection of adenovirus expressing HIF-2α (Ad-Epas1) or NAMPT (Ad-Nampt). The functions of SIRT in OA were examined by IA co-injection of SIRT inhibitors or adenovirus expressing individual SIRT isoforms or shRNA targeting specific SIRT isoforms. Results: HIF-2α activated the NAMPT-NAD<SUP>+</SUP>-SIRT axis in chondrocytes by upregulating NAMPT, which stimulated NAD<SUP>+</SUP> synthesis and thereby activated SIRT family members. The activated NAMPT-SIRT pathway, in turn, promoted HIF-2α protein stability by negatively regulating its hydroxylation and 26S proteasome-mediated degradation, resulting in increased HIF-2α transcriptional activity. Among SIRT family members (SIRT1-7), SIRT2 and SIRT4 were positively associated with HIF-2α stability and transcriptional activity in chondrocytes. This reciprocal regulation was required for the expression of catabolic matrix metalloproteinases (MMP3, MMP12, and MMP13) and OA cartilage destruction caused by IA injection of Ad-Epas1 Ad-Nampt. Conclusion: The reciprocal regulation of HIF-2α and the NAMPT-NAD<SUP>+</SUP>-SIRT axis in articular chondrocytes is involved in OA cartilage destruction caused by HIF-2α or NAMPT.

Role of interleukin-10 in endochondral bone formation

Kim, G.,Jung, Y.K.,Park, H.R.,Lee, E.J.,Choi, J.Y.,Beier, F.,Han, S.W. Published for the Society by Baillère Tinda 2013 Osteoarthritis and cartilage Vol.21 No.suppl

PCB126 induces apoptosis of chondrocytes via ROS-dependent pathways

Lee, H.G.,Yang, J.H. Published for the Society by Baillère Tinda 2012 OSTEOARTHRITIS AND CARTILAGE Vol.20 No.10

Objective: Chondrocyte apoptosis represents an important component in the osteoarthritis (OA) pathogenesis. This study sought to investigate the potential of polychlorinated biphenyl (PCB)126, the most potent and ubiquitous environmental pollutant of PCB congeners, on chondrocyte apoptosis and its mechanism of action. Methods: Rabbit articular chondrocytes cultured from tibial and femoral in cartilage were exposed to PCB126. Productions of reactive oxygen species (ROS) and nitric oxide (NO) and nuclear factor-kB (NF-kB) binding activity were measured. After 24 h exposure to PCB126, the apoptotic cell death was detected by caspase-3 activity, enzyme-linked immunosorbent assay (ELISA) using antibodies against DNA and histone, and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end-labeling (TUNEL) staining. Results: PCB126 generated ROS, which was blocked by the antioxidants (N-acetylcystein and trolox), or the aryl hydrocarbon receptor (AhR) inhibitor, α-naphthoflavone (α-NF). PCB126 exposure also increased NO production and NF-kB binding activity in the chondrocytes, which were blocked by the iNOS inhibitor, N-monomethyl-l-arginine (l-NMMA). All apoptosis detection techniques used in this study revealed an increase of apoptotic effects by PCB126 exposure, which was blocked by inhibitors of ROS or iNOS. This is the first report to demonstrate the potential of a PCB congener to induce chondrocytes apoptosis, which could be an initial process in cartilage degradation. Conclusions: PCB may be an initiator of chondrocyte apoptosis, which is closely linked to degradation of cartilage in OA pathogenesis. This study may contribute to identifying the possible causes of arthritis in our environment.

CXC chemokine ligand 12a enhances chondrocyte proliferation and maturation during endochondral bone formation

Kim, G.W.,Han, M.S.,Park, H.R.,Lee, E.J.,Jung, Y.K.,Usmani, S.E.,Ulici, V.,Han, S.W.,Beier, F. Published for the Society by Baillère Tinda 2015 OSTEOARTHRITIS AND CARTILAGE Vol.23 No.6

Objective: We investigated the roles of CXC chemokine ligand 12a (CXCL12a), also known as stromal cell-derived factor-1α (SDF-1α), in endochondral bone growth, which can give us important clues to understand the role of CXCL12a in osteoarthritis (OA). Methods: Primary chondrocytes and tibial explants from embryonic 15.5 day-old mice were cultured with recombinant mouse CXCL12a. To assess the role of CXCL12a in chondrogenic differentiation, we conducted mesenchymal cell micromass culture. Results: In tibia organ cultures, CXCL12a increased total bone length in a dose-dependent manner through proportional effects on cartilage and bone. In accordance with increased length, CXCL12a increased the protein level of proliferation markers, such as cyclin D1 and proliferating cell nuclear antigen (PCNA), in primary chondrocytes as well as in tibia organ culture. In addition, CXCL12a increased the expression of Runx2, Col10 and MMP13 in primary chondrocytes and tibia organ culture system, implying a role of CXCL12a in chondrocyte maturation. Micromass cultures of limb-bud mesenchymal progenitor cells (MPCs) revealed that CXCL12a has a limited effect on early chondrogenesis, but significantly promoted maturation of chondrocytes. CXCL12a induced the phosphorylation of p38 and Erk½ MAP kinases and IκB. The increased expression of cyclin D1 by CXCL12a was significantly attenuated by inhibitors of MEK1 and NF-κB. On the other hand, p38 and Erk½ MAP kinase and NF-κB signaling were associated with CXCL12a-induced expression of Runx2 and MMP13, the marker of chondrocyte maturation. Conclusion: CXCL12a promoted the proliferation and maturation of chondrocytes, which strongly suggest that CXCL12a may have a negative effect on articular cartilage and contribute to OA progression.

Ucma, a direct transcriptional target of Runx2 and Osterix, promotes osteoblast differentiation and nodule formation

Lee, Y.J.,Park, S.Y.,Lee, S.J.,Boo, Y.C.,Choi, J.Y.,Kim, J.E. Published for the Society by Baillère Tinda 2015 OSTEOARTHRITIS AND CARTILAGE Vol.23 No.8

Objective: Runt-related transcription factor 2 (Runx2) and Osterix (Osx) are the master transcription factors in bone formation. Nonetheless, genes acting downstream of both Runx2 and Osx have yet to be fully characterized. Here, we investigate the downstream targets of both Runx2 and Osx in osteoblasts. Materials and methods: DNA microarray analysis was conducted on calvarial RNA from wild-type, Runx2 heterozygous, Osx heterozygous, and Runx2/Osx double heterozygous embryos. Expression and transcriptional responses of the selected target gene were analyzed in MC3T3-E1 osteoblastic cells. Results: The expression of unique cartilage matrix-associated protein (Ucma) was decreased in Runx2/Osx double heterozygous embryos. In contrast, Ucma expression was increased in osteoblasts overexpressing both Runx2 and Osx. Ucma expression was initiated mid-way through osteoblast differentiation and continued throughout the differentiation process. Transcriptional activity of the Ucma promoter was increased upon transfection of the cells with both Runx2 and Osx. Runx2-and Osx-mediated activation of the Ucma promoter was directly regulated by Runx2-and/or Sp1-binding sites within its promoter. During osteoblast differentiation, the formation of mineralized nodules in Ucma-overexpressing stable clones occurred earlier and was more enhanced than that in the mock-transfected control. Mineralized nodule formation was strongly augmented in the cells cultured in a medium containing secretory Ucma proteins. Conclusion: Ucma is a novel downstream gene regulated by both Runx2 and Osx and it stimulates osteoblast differentiation and nodule formation.

Fully automated segmentation of cartilage from magnetic resonance images using improved 3D shape context and active shape model

Ye, T.,Cui, X.,Kim, H. Published for the Society by Baillère Tinda 2015 OSTEOARTHRITIS AND CARTILAGE Vol.23 No.suppl2

Polyethylene wear particles play a role in development of osteoarthritis via detrimental effects on cartilage, meniscus, and synovium

Park, D.Y.,Min, B.H.,Kim, D.W.,Song, B.R.,Kim, M.,Kim, Y.J. Published for the Society by Baillère Tinda 2013 Osteoarthritis and cartilage Vol.21 No.12

Objective: While ultra-high molecular weight polyethylene (UHMWPE) wear particles are known to cause periprosthetic osteolysis, its interaction with other intra-articular tissues in the case of partial joint arthroplasties is not well understood. We hypothesized that UHMWPE particles per se would interact with intra-articular tissue, which by acting as inflammatory reservoirs, would subsequently induce osteoarthritic (OA) changes. Our goal was to assess the inflammatory response, phagocytic activity, as well as apoptosis of intra-articular cells in the presence of UHMWPE particles in vitro, and the in vivo response of those tissues after intra-articular injection of particles in a murine model. Design: Three cell types were used for the in vitro study; chondrocytes, meniscal fibrochondrocytes, and synoviocytes. Each cell type was cultured with two different concentrations of UHMWPE particles. Pro-inflammatory cytokine production, phagocytosis, and apoptosis were analyzed. In vivo experiments were done by injecting two concentrations of UHMWPE particles into normal and murine OA model knee joints. Results: In vitro experiments showed that UHMWPE particles increase pro-inflammatory cytokine and mediator (IL-1β, IL-6, TNF-α, Nitric Oxide, and Prostaglandin E2) production, phagocytosis of particles, and apoptosis in all cell types. In vivo experiment showed degeneration of cartilage and meniscus, as well as synovitis after particle injection. Conclusions: UHMWPE wear particles per se exert detrimental effects in cartilage, synovium, and meniscus of the knee joint resulting in pro-inflammatory cytokine release, phagocytosis of particles and apoptosis. Particles induced and exacerbated OA changes in a murine model.

Stem cells for cartilage repair: what exactly were used for treatment, cultured adipose-derived stem cells or the unexpanded stromal vascular fraction?

Lee, J.I.,Balolong, E.,Han, Y.,Lee, S. Published for the Society by Baillère Tinda 2016 OSTEOARTHRITIS AND CARTILAGE Vol.24 No.7